31P Magnetic Resonance Spectroscopy (MRS) of tissues in vivo has revealed that phospho-monoesters (PMEs), in particular phosphorylcholine (PCho) and phosphorylethanolamine (PEth), are usually higher in proliferating tissues (e.g. tumors) than in non-proliferating normal tissues. There is considerable excitement about the use of these resonances as markers for tumor staging and therapeutic efficacy. Radiodiagnostic use of these resonances requires a more solid understanding of the factors and mechanisms regulating their metabolism. Most work to date has investigated these compounds vis-a-vis their roles in phospholipid synthesis. Consequently, phosphomonoesters themselves have not been greatly studied. In vitro, we and others have observed that the levels of PCho and PEth vary with culture density, perfusion and proliferation, suggesting that PME levels are influenced by both the extrinsic availability of substrates and by intrinsic differences between cell type and proliferative status. We hypothesize that the rate-limiting step(s) in PCho and PEth metabolism will vary between cell type and with proliferative status and that, in some cases more than one step may be involved (e.g. synthesis, diffusion, transport and phosphorylation of precursor amines, as well as the utilization of the PMEs in phospholipid synthesis). Defining the regulation of PME metabolism in a predictive model will allow PCho and PEth levels to be useful in determining the physiology and the environment of tumor cells in vivo. We propose to characterize the behavior and biochemistry of PCho and PEth in rat gliomas and human breast cancer cells grown in vitro and in vivo. These cell types were chosen because of their clinical importance and because previous work using these cell lines in our laboratory has indicated that they regulate their PME levels differently. Understanding the mechanism(s) behind these different phenotypes will help us understand the regulation of PME levels in vivo. The regulation of PME levels will be characterized using a combination of 31P MRS, 13C MRS and radiolabel techniques. 31P MRS will be used to assess the effects of exogenous amines and proliferation on steady-state PME levels, 13C MRS will identify the pathways of PME metabolism, and radiolabels will be used to quantify the kinetics of the PME-metabolizing pathways. These data will be used to identify the factors regulating PME metabolism and to develop a general model for PME metabolism in vivo. This project will test these models by comparing the behavior of PMEs in vitro to the same cells grown in vivo.